The long-term objectives of this proposal are to understand the way in which multipotent embryonic cells detect their position with respect to overall body pattern, and the way in which position- specific environmental cues act on the multipotent cell to commit it to one of its alternative developmental pathways. These problems will here e examined in the leech embryo, whose normal development is comprised of an invariant sequence of cell lineages in which certain uniquely identifiable cells choose between alternative pathways on the basis of position. One specific aim is to use single cell ablations to identify the cells which generate and the cells which receive positional cues. A second aim is to characterize the molecular basis of these positional cues. Such cues may be transmitted via gap junctions, and this hypothesis will be examined by injecting the interacting cells with antibodies that bind to the major gap junction protein in order to block transjunctional conductance. To generate effective antibodies, a gene for the leech gap junction protein is being isolated and sequenced, and portions of the predicted polypeptide will be used to immunize rabbits. The role of the ionic environment on the interaction of these embryonic cells will also be examined. A third aim is to identify molecular changes that occur in the multipotent cells in associated with its commitment to one of its alternative development pathways. Toward this end, a leech embryo cDNA library will be constructed and differential screening used as a means of identifying gene transcripts that are uniquely associated with a particular cell commitment. The final specific aim is to compare two segmentally homologous development pathways where similar cell commitments seem to the governed by different development mechanisms. The results of this study will lend insight into an important and poorly understood aspect of cell biology, i.e. the detailed mechanisms by which spatial patterns of cell differentiation arise during development, and are therefore relevant to the goals of human medicine.